Lateral-flow immunoassays, with their ease of use, speed and reliability, are widely used for self-testing and in the clinical setting. Lateral-flow immunoassays are probably the most common non-electrical method used in rapid medical diagnostics to detect the presence of a specific analyte in a liquid sample.
In the general method, a liquid sample suspected of containing the analyte is applied to a porous carrier. Different porous materials are commonly used for the porous carrier, and can differ in pore size, flow rate, protein-binding specifications and pre-treatment, etc. Essentially, all of the physical activities (e.g., liquid migration) and chemical reactions take place in the porous carrier, in the following order.
First a liquid sample to be tested is introduced to a designated area in the sampling-end (also referred as the “proximal end” or “wet end”) of the porous carrier, for a measured time e.g. 5 seconds or in a measured volume e.g. 2 drops. From this point forward, the liquid sample migrates within the porous carrier to the direction of the dry end (also referred as the “distal end”). At the outset of the migration, the liquid sample is frequently optimized for reaction by means of chemicals e.g. pH agents or buffers, surfactants, and/or blockers impregnated into the porous carrier.
Second, while migrating in the porous carrier, the sample mobilizes a labeled reagent that has been reversibly (temporarily) immobilized in the porous carrier. The zone where the mobilizable labeled reagent is located is often referred to as the “labeling zone”, but can be referred to as the “reversible immobilization zone” or “mobilization zone”—the terms are equivalent.
Third, while analyte is reacting with the mobilized labeled reagent, the liquid sample and mobilized labeled reagent migrates further within the porous carrier to the detection zone, (which may also be referred to the “irreversible immobilization zone” or merely the “immobilization zone”) where reagent that binds the same analyte is fixed or immobilized, usually in the form of a line. When analyte is present in the liquid sample, a “sandwich” in the form of the mobilized labeled reagent:analyte:immobilized reagent is formed, and the resulting concentration of the labeled reagent leads to a visible line appearing in the detection zone, which is indicative of a positive result.
Lastly, remaining sample liquid, together with the rest of the labeled reagent further migrates to a control zone, where a second line appears indicating that sample has progressed through the detection and control zones and that the assay has provided a valid test result. The rest of the sample and the remaining labeled reagent then migrate to a porous sink. Labeled reagent remaining in the porous carrier (other than in the detection zone, control zone or sink) makes up any background signal. In some instances where the migration direction reverses, so called “back flow,” occurs. Furthermore, the porous carrier can be pre-treated with chemicals e.g. surfactants.
Lateral-flow immunoassays can also function on the basis of competitive binding of the analyte. In these devices, lack of the test line generally indicates a positive result.
The most common example of a lateral-flow immunoassay device is a pregnancy test. These devices are commonly provided for home use, in a plastic housing with a fibrous or a porous extension, which can be held to a urine stream to collect urine sample into the housing. The urine sample collected this way then migrates to the porous carrier, which contains the labeled reagent and the series of events mentioned above starts. The analyte detected in a pregnancy test is Human Chorionic Gonadotropin (hCG) and the reagents commonly used are anti-hCG monoclonal or polyclonal antibodies. The most common labels are gold or latex particles.
Another known example of a lateral-flow immunoassay device commonly provided for home use is an ovulation test, the analyte being Luteinizing Hormone (LH) and reagents being anti-LH, and the rest of the device being similar to a pregnancy test.
A professional format of lateral-flow immunoassay devices commonly referred to as cassette tests, have smaller housings and a sample application orifice instead of the fibrous extension. The sample orifice exposes part of the porous carrier where a liquid sample can be dispensed with a pipette, directly to the porous carrier.
A low cost format of lateral-flow immunoassay devices is a dipstick test in the shape of a strip.
A diversity of test designs, against an ever increasing number of analytes, especially in the cassette and dipstick format, exists in the market indicating the wide acceptance of the method.